Impaired Gut-Liver-Brain Axis in Patients with Cirrhosis

Abstract

Cirrhosis is associated with brain dysfunction known as hepatic encephalopathy (HE). The mechanisms behind HE are unclear although hyperammonemia and systemic inflammation through gut dysbiosis have been proposed. We aimed to define the individual contribution of specific gut bacterial taxa towards astrocytic and neuronal changes in brain function using multi-modal MRI in patients with cirrhosis. 187 subjects (40 controls, 147 cirrhotic; 87 with HE) underwent systemic inflammatory assessment, cognitive testing, stool microbiota analysis and brain MRI analysis. MR spectroscopy (MRS) changes of increased Glutamate/glutamine, reduced myo-inositol and choline are hyperammonemia-associated astrocytic changes, while diffusion tensor imaging (DTI) demonstrates changes in neuronal integrity and edema. Linkages between cognition, MRI parameters and gut microbiota were compared between groups. We found that HE patients had a significantly worse cognitive performance, systemic inflammation, dysbiosis and hyperammonemia compared to controls and cirrhotics without HE. Specific microbial families (autochthonous taxa negatively and Enterobacteriaceae positively) correlated with MR spectroscopy and hyperammonemia-associated astrocytic changes. On the other hand Porphyromonadaceae, were only correlated with neuronal changes on DTI without linkages with ammonia. We conclude that specific gut microbial taxa are related to neuronal and astrocytic consequences of cirrhosis-associated brain dysfunction.

Figure 1

Microbiota analysis of stool 1(A). LEfSe predictions for bacterial families found in stool for healthy controls compared to cirrhosis. The histogram shows bacterial families in red bars that were significantly higher in control stool microbiota. LDA (linear discriminant analysis) score on the x-axis represents log changes in relative bacterial family representation in healthy controls compared to cirrhotic patients. The cladogram shows the phylogenetic relationship between the bacterial families that were higher in controls compared to cirrhotic patients that are represented in the red in the histogram. 1(B) LEfSe predictions for bacterial families found in stool for patients with hepatic encephalopathy. The histogram shows bacterial families in green bars that were significantly higher in cirrhotics with HE while those in red show bacterial families that were significantly higher in cirrhotics without HE. LDA (linear discriminant analysis) score on the x-axis represents log changes in relative bacterial family representation between the two groups. The cladogram shows the phylogenetic relationship between the bacterial families that were different in cirrhotics with HE compared to cirrhotics without HE representing the same colors for the groups as in the histogram.

Figure 2. Correlation networks of microbiota, cognitive tests and brain MRS values in controls.

Red nodes: stool bacterial families, light green: cognitive test results, dark green: brain MRS values, blue nodes: serum results. Red connections between nodes indicate negative correlation while blue connections are positive linkages. WM: right parietal white matter, PGM: posterior gray matter, ACC: anterior cingulate cortex, Cho: creatine ratio of choline, mI: creatine ratio of myoinositol, Glx: creatine ratio of glutamate + glutamine, BDT: block design test, DST: digit symbol test, NCT-A: number connection test-A, NCT-B: number connection test-B, Dotting: serial dotting test, LTT: line tracing test, targets and lures: outcomes of inhibitory control test. A high score on targets, digit symbol and block design indicates good cognitive performance while a high score on the rest of the cognitive tests suggests the opposite. Control network: There are few connections but the autochthonous family, Lachnospiraceae is positively linked with ACC mI and Alcaligenaceae, member of Proteobacteria is linked negatively to mI in PGM.

Figure 3. Correlation networks of microbiota, cognitive tests and brain MRS values in all cirrhotic patients.

Red nodes: stool bacterial families, light green: cognitive test results, dark green: brain MRS values, blue nodes: serum results. Red connections between nodes indicate negative correlation while blue connections are positive linkages. WM: right parietal white matter, PGM: posterior gray matter, ACC: anterior cingulate cortex, Cho: creatine ratio of choline, mI: creatine ratio of myoinositol, Glx: creatine ratio of glutamate + glutamine, BDT: block design test, DST: digit symbol test, NCT-A: number connection test-A, NCT-B: number connection test-B, Dotting: serial dotting test, LTT: line tracing test, targets and lures: outcomes of inhibitory control test. A high score on targets, digit symbol and block design indicates good cognitive performance while a high score on the rest of the cognitive tests suggests the opposite. Cirrhosis network: In all cirrhosis subjects, there were expected positive linkages between Glx, MELD and ammonia and negative between these and mI and Cho. These were related to poor cognitive performance. Autochthonous taxa (Lachnospiraceae, Ruminococcaeae and Incertae sedis XIV) were negatively correlated with MELD score and Glx while potentially pathogenic Enterococcaceae were positively related to MELD.

Figure 4. Correlation networks of microbiota, cognitive tests and brain MRS values in cirrhotic patients with HE.

Red nodes: stool bacterial families, light green: cognitive test results, dark green: brain MRS values, blue nodes: serum results. Red connections between nodes indicate negative correlation while blue connections are positive linkages. WM: right parietal white matter, PGM: posterior gray matter, ACC: anterior cingulate cortex, Cho: creatine ratio of choline, mI: creatine ratio of myoinositol, Glx: creatine ratio of glutamate + glutamine, BDT: block design test, DST: digit symbol test, NCT-A: number connection test-A, NCT-B: number connection test-B, Dotting: serial dotting test, LTT: line tracing test, targets and lures: outcomes of inhibitory control test. A high score on targets, digit symbol and block design indicates good cognitive performance while a high score on the rest of the cognitive tests suggests the opposite. In HE patients, there was evidence of a robust correlation network in which autochthonous bacterial families, Ruminococcaeae and Incertae sedis XIV were negatively correlated with MELD score and to MRS findings of increased Glx. Ammonia was again negatively related to good cognition and positively with Glx. Families such as Streptococcacae, Lactobacillaceae and Peptostreptococcaceae were related to neuro-inflammation and poor cognition.

Figure 5. Correlation networks of microbiota, cognitive tests and brain MRS values in cirrhotic patients without HE.

Red nodes: stool bacterial families, light green: cognitive test results, dark green: brain MRS values, blue nodes: serum results. Red connections between nodes indicate negative correlation while blue connections are positive linkages. WM: right parietal white matter, PGM: posterior gray matter, ACC: anterior cingulate cortex, Cho: creatine ratio of choline, mI: creatine ratio of myoinositol, Glx: creatine ratio of glutamate + glutamine, BDT: block design test, DST: digit symbol test, NCT-A: number connection test-A, NCT-B: number connection test-B, Dotting: serial dotting test, LTT: line tracing test, targets and lures: outcomes of inhibitory control test. A high score on targets, digit symbol and block design indicates good cognitive performance while a high score on the rest of the cognitive tests suggests the opposite. No-HE patients showed similar correlations between autochthonous taxa and good cognition and lesser neuro-inflammation while members of Proteobacteria such as Enterobacteriaceae and Pastuerellaceae were linked with worse neuro-inflammation as were Streptococcaceae.